Non-peptidic compounds are described for treatment of autoimmune diseases. Of particular interest are pipecolic acid derivatives compounds incorporating a characterizing segment condensed from pipecolic acid, aspartic acid, proline and threonine, or derivatives thereof, which compounds are useful to treat rheumatoid arthritis.
Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation of the synovium. This disease is triggered by an immune response generated via the molecular recognition of the T-cell receptor on CD4-positive T cells with a complex of disease-inducing peptides bound to Human Leukocyte Antigen (HLA) class II molecules.
Rheumatoid arthritis (RA) is associated with the expression of certain HLA class II molecules, particularly the DR4-dw4, as well as DR1 and DR4-dw14. It is known that blockade of the interaction between a given class II molecule, peptide ligand, and T cell receptor inhibits specific T cell responses both in vitro and in vivo. It is further known that blockade of the above interaction in animal models of autoimmunity prevents or ameliorates autoimmune disease. Inhibitor compounds which block the binding of disease-inducing peptides to an RA-associated HLA molecule, but which will not interfere with a patient""s ability to generate other class II-restricted immune responses, constitutes a selective immunosuppressive anti-RA therapy. Compounds which compete with disease-inducing endogenous peptides for binding to RA-associated HLA molecules and may thereby inhibit disease.
Other therapeutic strategies which are directed at the T cell, such as total lymphoid irradiation, thoracic duct drainage, cyclosporin A, anti-CD4 monoclonal antibody, and other monoclonal antibodies directed at T cell determinants, result in some cases in clinical improvement of rheumatoid arthritis, but these therapies are also associated with side effects. For instance, conventional general immunosuppressives increase the risk of opportunistic infections and cancer.
PCT App Pub #WO 93/05011, published Mar. 18, 1993, describes a series of oligopeptides, typically comprised of ten or more amino acids, which bind to HLA Class II molecules for indication against autoimmune diseases These larger peptides incorporating proteogenic amino acids can have poor oral bioavailability and poor plasma stability, as well as difficulty in penetrating cell membranes for proper tissue distribution.
A series of heptapeptides, such as recently reported to bind to HLA Class II molecules, similarly comprising natural, mammalian amino acids, unblocked at the C and N termini, are likely to be unstable in vivo and to possess poor bioavailability [Hammer et al, PNAS, 91, 4456-4460 (1994)].
Treatment of an autoimmune or inflammatory disease, or a hypersensitive reaction of the acute or delayed type, or an allergic reaction or asthmatic disorder, or treatment of dermatitits, arthritis, meningitis, granulomas, vasculitis or septic shock, may be accomplished by preventing or suppressing an immume response in a treatment subject by use of a pipecolic acid derivative of a proline threonine amide selected from a family of compounds of general Formula I: 
wherein R1 is selected from alkyl, phenyl, cycloalkyl rings having four to ten ring-member carbon atoms, bicycloalkyl fused ring systems having seven to nine ring-member carbon atoms, heteroaryl, heteroarylalkyl, benzo-fused-heteroaryl and benzo-fused-heteroarylalkyl wherein said heteroaryl moiety or fragment is a 5- or 6-ring-member fully-unsaturated ring system having one hetero atom as a ring member, said hetero atom selected from oxygen, nitrogen and sulfur atoms, and wherein any of said heteroaryl, heteroarylalkyl, benzo-fused-heteroaryl and benzo-fused-heteroarylalkyl may be attached to the nucleus of Formula I as an R1 substituent through a bond formed at any said ring-member atom or any atom of the alkyl portion of said R1 substituent where said bond is capable of forming a stable compound;
wherein R2 is selected from hydrido, lower alkyl, cyclohexyl and phenyl;
wherein R3 is selected from hydrido, hydroxy, lower alkyl, phenyl, acetyl(Lys)NHxe2x80x94, acetyl(Tyr)NHxe2x80x94, acetyl(Thr)NHxe2x80x94, acetylamino, propionylamino and benzyloxycarbonylamino;
wherein R4 is selected from hydrido, lower alkyl and phenyl;
wherein R5 is selected from hydrido, lower alkyl, phenyl, benzyl, hydroxyphenyl, hydroxybenzyl, aminoalkyl, mono-alkyl-substituted-aminoalkyl and radicals provided by B-Het-A;
wherein Het is selected from heteroaryl moieties consisting of monocyclic and fused bicyclic ring systems having a total of five to fourteen ring members and with one to six ring members being selected from hetero atoms provided by oxygen, nitrogen and sulfur atoms, wherein said monocyclic ring system and at least one ring system of said fused bicyclic ring system is fully unsaturated, and
wherein Het is further selected from heterocyclic moieties consisting of monocyclic and fused polycyclic ring systems having a total of four to twelve ring members and with one to six ring members selected from hetero atoms provided by oxygen, nitrogen and sulfur atoms, wherein said monocyclic ring-system and at least one ring system of said fused polycyclic ring system is fully saturated or partially unsaturated,
wherein A is a single covalent bond or is a divalent radical selected from 
wherein R13 is lower alkyl;
wherein B is one or more substituents attached at a substitutable position on Het of Het-A, said substituent selected from hydrido, hydroxy, alkyl, cycloalkyl, cycloalkylalkyl, alkoxy, hydroxyalkyl, alkoxyalkyl, carboxy, alkenyl, alkynyl, halo, haloalkyl, oxo, cyano, benzyl and phenyl;
wherein R6 is selected from hydrido, lower alkyl, hydroxy, alkoxy, alkoxyalkyl, carboxyalkyl, alkoxycarbonyl, alkoxycarbonyloxy, aminoalkyl, mono-alkyl-substituted-aminoalkyl, amido and amidoalkyl;
wherein R7 is selected from carboxyl, lower alkyl, amido and methylthiomethyl;
wherein R8 is selected from hydrido, methyl and ethyl;
wherein R9 is selected from hydrido, lower alkyl, alkoxy and phenyl;
wherein R10 is hydrido or hydroxy;
wherein R11 is hydrido or methyl;
wherein R12 is selected from lower alkyl, phenyl, phenylalkyl, cycloalkyl, cycloalkylalkyl, 
wherein each of R14 through R17 is independently selected from hydrido, hydroxy, alkyl, hydroxyalkyl, alkoxy, alkoxyalkyl, cycloalkyl, cycloalkylalkyl, halo, haloalkyl, cyano, benzyl and phenyl;
wherein each of R18 and R19 is independently selected from hydrido, alkyl, cycloalkyl, cycloalkylalkyl, benzyl and phenyl;
or a pharmaceutically-acceptable amide, ester or salt thereof.
Examples of heteroaryl 5- or 6-ring member monocyclic ring systems and benzo-fused bicyclic ring systems, having one hetero atom as a ring member selected from oxygen, nitrogen and sulfur atoms, and which ring systems are fully unsaturated, i.e. xe2x80x9caromaticxe2x80x9d in character, in at least one ring, are as follow: 
Examples of heterocyclic-type monocyclic or polycyclic ring systems having four to twelve ring members with one to six hetero atoms as ring members, said hetero atoms selected from oxygen, nitrogen and sulfur atoms, and wherein at least one of such ring systems is fully saturated or partially unsaturated, and wherein the polycyclic ring system may be composed of a benzo-fused ring, are as follow: 
Examples of heteroaryl monocyclic and bicyclic ring systems and benzo-fused polycyclic ring systems, having one to six hetero atoms as ring members selected from oxygen, nitrogen and sulfur atoms, and which ring systems are fully unsaturated, i.e. xe2x80x9caromaticxe2x80x9d in character, in at least one of the ring system, are as follow: 
A preferred family of compounds consists of compounds of Formula I:
wherein R1 is selected from cyclopentyl, cyclohexyl, cycloheptyl, norbornanyl, phenyl, furyl, pyrrolyl, thienyl, chromanyl, isochromanyl, benzothienyl, pyridyl, indolizinyl, isoindolyl, indolyl, 3H-indolyl, quinolizinyl, quinolyl, isoquinolyl, azetidinyl, thioazetidinyl, pyrrolidinyl, pyrrolinyl, oxazolidinyl,thiazolidinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidinyl, piperazinyl, 1,3-morpholino, 1,4-morpholino, 1,4-thiomorpholino, azepinyl, oxazopinyl, thiazopinyl, oxazocinyl, thiazocinyl, azoninyl, oxazabicyclo, benzo-fused-oxazolidinyl, benzo-fused-thiazolidinyl, benzo-fused-morpholino, benzo-fused thiomorpholinyl, benzo-fused-thiazopinyl, benzo-fused oxazopinyl, benzo-fused-oxazocinyl, benzo-fused-oxazoninyl, tropanyl and benzo-fused-oxazobicyclo;
wherein R2 is selected from hydrido, methyl, ethyl, propyl, cyclohexyl and phenyl;
wherein R3 is selected from hydrido, hydroxy, methyl, ethyl, phenyl, acetyl(Lys)NHxe2x80x94, acetyl(Tyr)NHxe2x80x94, acetyl(Thr)NHxe2x80x94, acetylamino, propionylamino and benzyloxycarbonylamino;
wherein R4 is hydrido or methyl;
wherein R5 is selected from hydrido, n-propyl, isopropyl, n-butyl, isobutyl, phenyl, benzyl, hydroxyphenyl, hydroxybenzyl, aminopropyl, aminobutyl and radicals provided by B-Het-R13 and 
wherein Het is selected from furyl, pyrrolyl, thienyl, chromanyl, isochromanyl, benzothienyl, pyridyl, indolizinyl, isoindolyl, indolyl, 3H-indolyl, quinolizinyl, quinolyl, isoquinolyl, imidazolyl, pyrazolyl, oxazolidyl, thiazolidyl, isothiazolidyl, isoxazolidyl, furazanyl, pyrazinyl, pyrimidinyl, pyridazinyl, indazolyl, purinyl, phthalazinyl, naphthyridinyl, quinoxalinyl, quinazolinyl, cinnolinyl, pteridinyl, thieno-furanyl, furopyranyl, pyrido-oxazinyl, pyrazolo-oxazolyl, imidazo-thiazolyl, pyrazino-pyridazinyl, imidazo-thiazolyl, oxothiolo-pyrrolyl, imidazo-triazinyl, benzoxazinyl, azetidinyl, thioazetidinyl, pyrrolidinyl, pyrrolinyl, oxazolidinyl, thiazolidinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidinyl, piperazinyl, 1, 3-morpholino, 1,4-morpholino, 1,4-thiomorpholino, azepinyl, oxazopinyl, thiazopinyl, oxazocinyl, thiazocinyl, azoninyl, oxazabicyclo, benzo-fused-oxazolidinyl, benzo-fused-thiazolidinyl, benzo-fused-morpholino, benzo-fused thiomorpholinyl, benzo-fused-thiazopinyl, benzo-fused oxazopinyl, benzo-fused-oxazocinyl, benzo-fused-oxazoninyl, tropanyl and benzo-fused-oxazobicyclo;
wherein R13 is lower alkyl;
wherein B is one or more substituents attached at a substitutable position on Het, said substituent selected from hydrido, hydroxy, alkyl, cycloalkyl, cycloalkylalkyl, alkoxy, hydroxyalkyl, alkoxyalkyl, oxo, benzyl and phenyl;
wherein R6 is selected from hydrido, lower alkyl, hydroxy, methoxy carboxyalkyl, alkoxycarbonyl, alkoxycarbonyloxy, aminoalkyl, mono-alkyl-substituted-aminoalkyl, amido and amidoalkyl;
wherein R7 is selected from carboxyl, lower alkyl, amido and methylthiomethyl;
wherein R8 is hydrido or methyl;
wherein R9 is selected from hydrido, lower alkyl, methoxy and phenyl;
wherein R10 is hydrido or hydroxy;
wherein R11 is hydrido or methyl;
wherein R12 is selected from lower alkyl, phenyl, benzyl, phenylethyl, cyclohexylethyl, 
wherein each of R14 through R17 is independently selected from hydrido, hydroxy and alkyl;
or a pharmaceutically-acceptable amide, ester or salt thereof.
A more preferred family of compounds consists of compounds of Formula I:
wherein R1 is selected from cyclopentyl, cyclohexyl, cycloheptyl, norbornanyl, phenyl, azetidinyl, thioazetidinyl, pyrrolidinyl, pyrrolinyl, oxazolidinyl, thiazolidinyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidinyl, piperazinyl 1,3-morpholino, 1,4-morpholino, 1,4-thiomorpholino, azepinyl, oxazopinyl, thiazopinyl, oxazocinyl, thiazocinyl, azoninyl, oxazabicyclo and tropanyl;
wherein R2 is selected from hydrido, methyl, ethyl, propyl, acetyl(Lys)NHxe2x80x94, acetyl(Tyr)NHxe2x80x94, acetyl(Thr)NHxe2x80x94, cyclohexyl and phenyl;
wherein R3 is selected from hydrido, hydroxy, methyl, ethyl, phenyl, acetylamino, propionylamino and benzyloxycarbonylamino;
wherein R4 is hydrido or methyl;
wherein R5 is selected from hydrido, n-propyl, isopropyl, n-butyl, isobutyl, aminopropyl, aminobutyl, phenyl, hydroxyphenyl, benzyl, hydroxybenzyl and radicals provided by 
wherein Het is selected from azetidinyl, pyridinyl, isoindolyl, oxazolyl, isoxazolyl, indolyl, quinolyl, isoquinolyl, azetidinyl, thioazetidinyl, pyrrolidinyl, pyrrolinyl, oxazolidinyl, thiazolidinyl, imidazolyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidinyl, piperazinyl, 1,3-morpholino, 1,4-morpholino, 1,4-thiomorpholino, azepinyl, oxazopinyl, thiazopinyl, oxazocinyl, thiazocinyl, azoninyl, oxazabicyclo and tropanyl;
wherein R13 is selected from methyl, ethyl and propyl;
wherein B is one or more substituents attached at a substitutable position on Het, said substituent selected from hydrido, hydroxy, methyl, ethyl, propyl, oxo, benzyl and phenyl;
wherein R6 is selected from hydrido, methyl, hydroxy, methoxy, phenyl, alkoxycarbonyl, alkoxycarbonyloxy, aminoalkyl, mono-amido and amidoalkyl;
wherein R7 is selected from carboxyl, n-propyl, n-butyl, amido and methylthiomethyl;
wherein R8 is hydrido or methyl;
wherein R9 is selected from hydrido, lower alkyl, methoxy and phenyl;
wherein R10 is hydroxy;
wherein R11 is hydrido or methyl;
wherein R12 is selected from lower alkyl, phenyl, phenylethyl, cyclohexylethyl, 
wherein each of R14 through R17 is independently selected from hydrido, hydroxy, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, benzyl and phenyl;
or a pharmaceutically-acceptable amide, ester or salt thereof.
A highly preferred family of compounds consists of compounds of Formula I:
wherein R1 is phenyl or cyclohexyl; wherein R2 is hydrido or methyl; wherein R3 is selected from hydrido, hydroxy, acetyl (Lys)NHxe2x80x94, acetyl(Tyr)NHxe2x80x94, acetyl(Thr)NHxe2x80x94, acetylamino, propionylamino and benzyloxycarbonylamino; wherein R4 is hydrido; wherein R5 is selected from isopropyl, isobutyl, n-propyl, n-butyl, aminopropyl, aminobutyl, phenyl, benzyl, para-hydroxyphenyl, para-hydroxybenzyl, imidazolcarbonylethyl, imidazolcarbonylpropyl, pyrrolidinylcarbonylethyl, pyrrolidinylcarbonylpropyl, azetidinylcarbonylethyl, azetidinylcarbonylpropyl, morpholinocarbonylethyl, morpholinocarbonylpropyl, piperazinocarbonylethyl, piperazinocarbonylpropyl, pyridinylcarbonylethyl, pyridinylcarbonylpropyl, oxazolylcarbonylethyl, oxazolylcarbonylpropyl, isoxazolylcarbonylethyl, isoxazolylcarbonylpropyl, azepinylcarbonylethyl and azepinylcarbonylethyl; wherein R6 is selected from hydrido, methyl, hydroxy, methoxy, phenyl and aminocarbonyl; wherein R7 is carboxyl or methylthiomethyl; wherein R8 is hydrido; wherein R9 is selected from hydrido, hydroxy, methyl, methoxy and phenyl; wherein R10 is hydroxy; wherein R11 is methyl; wherein R12 is selected from methyl, ethyl, propyl, butyl, isobutyl, xe2x80x94CH(iBu)CH2OH and xe2x80x94CH(iBu)CONH2; or a pharmaceutically-acceptable amide, ester or salt thereof.
The term xe2x80x9chydridoxe2x80x9d denotes a single hydrogen atom (H). This hydrido group may be attached, for example, to an oxygen atom to form a hydroxyl group; or, as another example, one hydrido group may be attached to a carbon atom to form a 
group; or, as another example, two hydrido atoms may be attached to a carbon atom to form a xe2x80x94CH2xe2x80x94 group. Where the term xe2x80x9calkylxe2x80x9d is used, either alone or within other terms such as xe2x80x9chaloalkylxe2x80x9d and xe2x80x9chydroxyalkylxe2x80x9d, the term xe2x80x9calkylxe2x80x9d embraces linear or branched radicals having one to about twenty carbon atoms or, preferably, one to about twelve carbon atoms. More preferred alkyl radicals are xe2x80x9clower alkylxe2x80x9d, radicals having one to about ten carbon atoms. Most preferred are lower alkyl radicals having one to about five carbon atoms. The term xe2x80x9ccycloalkylxe2x80x9d embraces cyclic radicals having three to about ten ring carbon atoms, preferably three to about six carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. The term xe2x80x9chaloalkylxe2x80x9d embraces radicals wherein any one or more of the alkyl carbon atoms is substituted with one or more halo groups, preferably selected from bromo, chloro and fluoro. Specifically embraced by the term xe2x80x9chaloalkylxe2x80x9d are monohaloalkyl, dihaloalkyl and polyhaloalkyl groups. A monohaloalkyl group, for example, may have either a bromo, a chloro, or a fluoro atom within the group. Dihaloalkyl and polyhaloalkyl groups may be substituted with two or more of the same halo groups, or may have a combination of different halo groups. A dihaloalkyl group, for example, may have two fluoro atoms, such as difluoromethyl and difluorobutyl groups, or two chloro atoms, such as a dichloromethyl group, or one fluoro atom and one chloro atom, such as a fluoro-chloromethyl group. Examples of a polyhaloalkyl are trifluoromethyl, 1,1-difluoroethyl, 2,2,2-trifluoroethyl, perfluoroethyl and 2,2,3,3-tetrafluoropropyl groups. The term xe2x80x9cdifluoroalkylxe2x80x9d embraces alkyl groups having two fluoro atoms substituted on any one or two of the alkyl group carbon atoms. The terms xe2x80x9calkylolxe2x80x9d and xe2x80x9chydroxyalkylxe2x80x9d embrace linear or branched alkyl groups having one to about ten carbon atoms any one of which may be substituted with one or more hydroxyl groups. The term xe2x80x9calkenylxe2x80x9d embraces linear or branched radicals having two to about twenty carbon atoms, preferably three to about ten carbon atoms, and containing at least one carbon-carbon double bond, which carbon-carbon double bond may have either cis or trans geometry within the alkenyl moiety. The term xe2x80x9calkynylxe2x80x9d embraces linear or branched radicals having two to about twenty carbon atoms, preferably two to about ten carbon atoms, and containing at least one carbon-carbon triple bond. The term xe2x80x9ccycloalkenylxe2x80x9d embraces cyclic radicals having three to about ten ring carbon atoms including one or more double bonds involving adjacent ring carbons. The terms xe2x80x9calkoxyxe2x80x9d and xe2x80x9calkoxyalkylxe2x80x9d embrace linear or branched oxy-containing radicals each having alkyl portions of one to about ten carbon atoms, such as methoxy group. The term xe2x80x9calkoxyalkylxe2x80x9d also embraces alkyl radicals having two or more alkoxy groups attached to the alkyl radical, that is, to form monoalkoxyalkyl and dialkoxyalkyl groups. The xe2x80x9calkoxyxe2x80x9d or xe2x80x9calkoxyalkylxe2x80x9d radicals may be further substituted with one or more halo atoms, such as fluoro, chloro or bromo, to provide haloalkoxy or haloalkoxyalkyl groups. The term xe2x80x9calkylthioxe2x80x9d embraces radicals containing a linear or branched alkyl group, of one to about ten carbon atoms attached to a divalent sulfur atom, such as a methythio group. Preferred aryl groups are those consisting of one, two, or three benzene rings. The term xe2x80x9carylxe2x80x9d embraces aromatic radicals such as phenyl, naphthyl and biphenyl. The term xe2x80x9caralkyl embraces aryl-substituted alkyl radicals such as benzyl, diphenylmethyl, triphenylmethyl, phenyl-ethyl, phenylbutyl and diphenylethyl. The terms xe2x80x9cbenzylxe2x80x9d and xe2x80x9cphenylmethylxe2x80x9d are interchangeable. The terms xe2x80x9cphenalkylxe2x80x9d and xe2x80x9cphenylalkylxe2x80x9d are interchangeable. An example of xe2x80x9cphenalkylxe2x80x9d is xe2x80x9cphenethylxe2x80x9d which is interchangeable with xe2x80x9cphenylethylxe2x80x9d. The terms xe2x80x9calkylarylxe2x80x9d, xe2x80x9calkoxyarylxe2x80x9d and xe2x80x9chaloarylxe2x80x9d denote, respectively, the substitution of one or more xe2x80x9calkylxe2x80x9d, xe2x80x9calkoxyxe2x80x9d and xe2x80x9chaloxe2x80x9d groups, respectively, substituted on an xe2x80x9carylxe2x80x9d nucleus, such as a phenyl moiety, which is then attached to the structure of Formula I or Formula II. The terms xe2x80x9caryloxyxe2x80x9d and xe2x80x9carylthioxe2x80x9d denote radicals respectively, provided by aryl groups having an oxygen or sulfur atom through which the radical is attached to a nucleus, examples of which are phenoxy and phenylthio. The terms xe2x80x9csulfinylxe2x80x9d and xe2x80x9csulfonylxe2x80x9d, whether used alone or linked to other terms, denotes, respectively, divalent radicals SO and SO2. The term xe2x80x9caralkoxyxe2x80x9d, alone or within another term, embraces an aryl group attached to an alkoxy group to form, for example, benzyloxy. The term xe2x80x9cacylxe2x80x9d whether used alone, or within a term such as acyloxy, denotes a radical provided by the residue after removal of hydroxyl from an organic acid, examples of such radical being acetyl and benzoyl. xe2x80x9cLower alkanoylxe2x80x9d is an example of a more prefered sub-class of acyl. The term xe2x80x9camidoxe2x80x9d denotes a radical consisting of nitrogen atom attached to a carbonyl group, which radical may be further substituted in the manner described herein. The amido radical can be attached to the nucleus of a compound of Formula I or Formula II through the carbonyl moiety or through the nitrogen atom of the amido radical. The term monoalkylaminocarbonylxe2x80x9d is interchangeable with xe2x80x9cN-alkylamidoxe2x80x9d. The term xe2x80x9cdialkylaminocarbonylxe2x80x9d is interchangeable with xe2x80x9cN,N-dialkylamidoxe2x80x9d. The term xe2x80x9calkenylalkylxe2x80x9d denotes a radical having a double-bond unsaturation site between two carbons, and which radical may consist of only two carbons or may be further substituted with alkyl groups which may optionally contain additional double-bond unsaturation. The term xe2x80x9cheteroarylxe2x80x9d, where not otherwised defined before, embraces aromatic ring systems containing one or two hetero atoms selected from oxygen, nitrogen and sulfur in a ring system having five or six ring members, examples of which are thienyl, furanyl, pyridinyl, thiazolyl, pyrimidyl and isoxazolyl. Such heteroaryl may be attached as a substituent through a carbon atom of the heteroaryl ring system, or may be attached through a carbon atom of a moiety substituted on a heteroaryl ring-member carbon atom, for example, through the methylene substituent of imidazolemethyl moiety. Also, such heteroaryl may be attached through a ring nitrogen atom as long as aromaticity of the heteroaryl moiety is preserved after attachment. For any of the foregoing defined radicals, preferred radicals are those containing from one to about ten carbon atoms.
Specific examples of alkyl groups are methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, methylbutyl, dimethylbutyl and neopentyl. Typical alkenyl and alkynyl groups may have one unsaturated bond, such as an allyl group, or may have a plurality of unsaturated bonds, with such plurality of bonds either adjacent, such as allene-type structures, or in conjugation, or separated by several saturated carbons.
Also included in the combination of the invention are the isomeric forms of the above-described compounds of Formula I, including diastereoisomers, regioisomers and the pharmaceutically-acceptable salts thereof. The term xe2x80x9cpharmaceutically-acceptable saltsxe2x80x9d embraces salts commonly used to form alkali metal salts and to form addition salts of free acids or free bases. The nature of the salt is not critical, provided that it is pharmaceutically-acceptable. Suitable pharmaceutically-acceptable acid addition salts may be prepared from an inorganic acid or from an organic acid. Examples of such inorganic acids are hydrochloric, hydrobromic, hydroiodic, nitric, carbonic, sulfuric and phosphoric acid. Appropriate organic acids may be selected from aliphatic, cycloaliphatic, aromatic, araliphatic, heterocyclic, carboxylic and sulfonic classes of organic acids, example of which are formic, acetic, propionic, succinic, glycolic, gluconic, lactic, malic, tartaric, citric, ascorbic, glucuronic, maleic, fumaric, pyruvic, aspartic, glutamic, benzoic, anthranilic, p-hydroxybenzoic, salicyclic, phenylacetic, mandelic, embonic (pamoic), methansulfonic, ethanesulfonic, 2-hydroxyethanesulfonic, pantothenic, benzenesulfonic, toluenesulfonic, sulfanilic, mesylic, cyclohexylaminosulfonic, stearic, algenic, xcex2-hydroxybutyric, malonic, galactaric and galacturonic acid. Suitable pharmaceutically-acceptable base addition salts include metallic salts made from aluminium, calcium, lithium, magnesium, potassium, sodium and zinc or organic salts made from N,Nxe2x80x2-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N-methylgluca-mine) and procaine All of these salts may be prepared by conventional means from the corresponding compound by reacting, for example, the appropriate acid or base with such compound.
Nomenclature used to define the amino acids used to make compounds of Formula I is that specified by the IUPAC [published in European Journal of Biochemistry, 138, 9-37 (1984)], wherein conventional representation of the peptides stipulates that in a peptide sequence the amino group appears to the left and the carboxyl group to the right. When the amino acid has enantiomeric forms, it is the L form of the amino acid which is represented unless otherwise stated. In the amino acid structural formulas, each residue is generally represented by a single or 3-letter designation, corresponding to the trivial name of the amino acid in accordance with the following list:
Another name for norvaline in n-propylglycine. The group 125I-Tyr indicates a radioactive mono-iodinated tyrosine residue.
CBA=3-cyclohexylbutyric acid
CPA=3-cyclohexylpropionic acid
Ac=acetyl
OBn=benzyloxy
Pec=pipecolic acid
Z=benzyloxycarbonyl
Compounds of the present invention function as HLA xe2x80x9cgroove blockersxe2x80x9d, referring to the peptide binding pocket or xe2x80x9cgroovexe2x80x9d present in the HLA, particularly the PR4-dw4, as well as DR1 and DR4-dw14. Because the compounds of the present invention are targeted to specific subsets of Class II molecules, greater specificity and better side-effect profiles are gained. Compounds of the present invention find utility in the treatment of other autoimmune diseases, including rheumatoid arthritis.
Thus, compounds of Formula I would be useful in suppressing immune response in a human or animal subject susceptible to or afflicted with an autoimmune disease or inflammatory disease. Examples of such treatable disease are systemic lupus erythematosis, multiple sclerosis, myesthenia gravis, thyroiditis, Grave""s disease, autoimmune hemolytic anemia, autoimmune hemolytic anemia, autoimmune thrombocytopenia purpura, mixed connective tissue disease, idiopathic Addison""s disease, Sjogren""s syndrome, insulin dependent diabetes mellitus, rheumatoid arthritis, psoriasis, glomerulonephritis, inflammatory bowel disease and Crohn""s Disease.
Compounds of Formula I would also be useful in suppressing immune response in a human or animal subject susceptible to or afflicted with an allergy, such as an asthmatic condition or reaction, urticaria or with airway hypersensitivity.
Compounds of Formula I would also be useful in suppressing immune response in a human or animal subject afflicted with or susceptible to septic shock.
Compounds of Formula I would also be useful in preventing or suppressing acute or delayed-type hypersensitivity responses or conditions resulting from or associated with hypersensitivity responses such as contact dermatitis, hemolytic anemias, antibody-induced thrombocytopenia, Goodpasture""s syndrome, hypersensitivity, pneumonitis, glomerulonephritis, granulomas, thyroiditis, encephelomyelitis and meningitis.
Of particular interest is use of a compound of Formula I to treat rheumatoid arthritis.
Compounds of Formula I would also be useful in adjunct therapy involving, typically, coadministration of a second immunosuppressive compound of Formula I or coadministration of an immunosuppressive agent of a different class of compounds. Such coadministration may provide a synergistic result between a compound of Formula I and an agent selected from one or more other classes of immunosuppressants. Such synergistic result allows for a lower dosage of another immunosuppressant agent having a toxic effect, such as a cyclosporin agent or steroid agent, such as cortisone and cortisol. Thus, use of a compound of Formula I in such synergistic combination allows utilization of the immunosuppressant benefits of a cyclosporin agent or steroid agent without, or with less of, the deleterious side effects of such agent. Examples of other immunosuppressant agents include a cyclosporin compound, Fujisawa FK-506 (macrolide lactone) compound, rapamycin, a glucocorticoid, an antiproliferative agent, a monoclonal antibody such as an anti-CD3 (anti-T cell receptor antibody), anti-CD5/CD7, anti-CD4 agent, an anti-IL-2 receptor (anti-cytokine receptor antibody) agent, an anti-IL-2 (anti-cytokine antibody), Nippon NKT-01 (15-deoxyspergualin) and Syntex RS-61443.

The compounds of Formula I can be prepared by coupling individual protected amino acid building blocks by standard solution or solid phase methods. The coupling reagents may be selected from those amide-forming reagents known in the art, such as carbodiimides, mixed carbonic anhydrides and active esters, but are not limited to these methods. The following Steps and Examples constitute specific exemplification of methods to prepare starting materials, intermediates and final compounds of Formula I. Those skilled in the art will readily understand that known variations of the conditions and processes of the following preparative procedures can be used to prepare the compounds of the Steps and Examples. All temperatures expressed are in degrees Centigrade.
Commercially available Boc-L-Thr(OBn)-OH (5 g) was mixed with 1-hydroxybenzotriazole (2.62 g), diisopropylethylamine (2.08 g) in methylene chloride (14 mL) and cooled to 0xc2x0 C. To this was added 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, followed by n-propylamine (1.14 g) 2 minutes later. After allowing the reaction mixture to stir at 0 for 5 hours, the organic solution was washed sequentially with water, 1N hydrochloric acid, 5% sodium bicarbonate, then dried (magnesium sulfate), filtered and evaporated to obtain an oily residue. This residue was taken up in 4M hydrogen chloride in dioxane solution (50 mL) and allowed to stand at room temperature for 30 minutes. The solvent was then evaporated and the product triturated with ether. The resulting solid was collected a filter plate. A portion of this material (1.5 g) was then mixed with Boc-L-Pro-OSu (1.5 g) in methylene chloride (20 mL), cooled to 0xc2x0 C., and diisopropylethylamine (675 mg) was added. After allowing the reaction mixture to stir at 0xc2x0 C. for 5 hours, the organic solution was washed sequentially with water, 1N hydrochloric acid, 5% sodium bicarbonate, then dried (magnesium sulfate), filtered and evaporated to obtain an oily residue. This residue was taken up in 4 M hydrogen chloride in dioxane solution (50 mL) and allowed to stand at room temperature for 30 minutes. The solvent was then evaporated and the product triturated with ether. The resulting solid was collected a filter plate to yield the title compound (1.45 g). 1H NMR (400 MHz): consistent with proposed structure.
Commercially available Boc-L-Asp(OBn)-OH (1 g) was mixed with 1-hydroxybenzotriazole (500 mg), the title compound from Step 1 (1 g) and diisopropylethylamine (736 mg) in methylene chloride (20 mL), and the solution was cooled to 0xc2x0 C. To this was added 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (710 mg) in one portion. After allowing the reaction mixture to stir at 0xc2x0 C. for 5 hours, the organic solution was washed sequentially with water, 1 N hydrochloric acid, 5% sodium bicarbonate, then dried (magnesium sulfate), filtered and evaporated to obtain an oily residue. This residue was taken up in 4 M hydrogen chloride in dioxane solution (50 mL) and allowed to stand at room temperature for 30 minutes. The solvent was then evaporated and the product triturated with ether. The resulting white solid was collected a filter plate to yield the title compound. 1H NMR (400 MHz): consistent with proposed structure.
Commercially available Boc-L-pipecolic acid (Boc-Pec, 100 mg) was mixed with 1-hydroxybenzotriazole (71 mg), the title compound from Step 2 (259 mg) and diisopropylethylamine (113 mg) in methylene chloride (2 mL), and the solution was cooled to 0xc2x0 C. To this was added 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (100 mg) in one portion. After allowing the reaction mixture to stir at 0xc2x0 C. for 5 hours, the organic solution was washed sequentially with water, 1N hydrochloric acid, 5% sodium bicarbonate, then dried (magnesium sulfate), filtered and evaporated. The residue was taken up in 4M hydrogen chloride in dioxane solution (50 mL) and allowed to stand at room temperature for 30 minutes. The solvent was then evaporated and the product triturated with ether. The resulting white solid was collected a filter plate to yield the title compound (223 mg). 1H NMR (400 MHz): consistent with proposed structure.
The same procedure as described in Step 3 was used to couple Boc-L-Lys(Z)-OH to the title compound of Step 3, yielding the title compound as a white solid (1.12 g)
Boc-Val-OH was dissolved in CH2Cl2, and cooled to 0xc2x0 C. via ice water bath. The reaction solution was treated with 1.2 eq of 1-hydroxybenzotriazole, 2 eq of diisopropylethylamine, and 1.5 eq of Pec-OMe.HCl. After all solids were dissolved, the reaction was treated with 1.2 eq of 1-(3-dimethyl-aminopropyl)-3-ethylcarbodiimide.HCl. The reaction flask was stoppered, sealed with parafilm, and placed in the refrigerator overnight. The CH2Cl2 was evaporated and replaced with EA. The organic solution was washed with water, 1.5 M HCl, and 5% NaHCO3, dried over Na2SO4, and thoroughly evaporated to a clear, colorless oil. The entire sample was then dissolved in 4.0M HCl in dioxane, stirred for one hour at rt, and thoroughly evaporated to yield Val-Pec-OMe.HCl as a white foam.
Commercially available (S)-3-phenylbutyric acid treated with rhodium-on-carbon in methanol at 60 psi and 60xc2x0 C. for 3.2 hours to yield (S)-3-cyclohexylbutyric acid. This acid was then converted to the acid chloride by treatment with 1 equivalent of oxalyl chloride and a catalytic amount of DMF, in toluene for 90 minutes. 250 mg of resulting (S)-3-cyclohexylbutyroyl chloride was dissolved in 10 mL ethyl acetate, and treated with 1.5 equivalents of the dipeptide, Val-Pec-OMe hydrochloride, followed by 3 equivalents of sodium bicarbonate dissolved in 2 mL of water. The reaction mixture stirred vigorously for 2 hours. The organic solution was then washed with 1.5 M HCl, dried over sodium sulfate, and evaporated to obtain a clear, colorless oil. This product was then treated with 3 equivalents of KOH dissolved in water and tetrahydrofuran and the reaction stirred vigorously for 12 hours. The tetrahydrofuran was evaporated, and the acid product was precipitated by acidifying the aqueous solution with conc. HCl. The mixture was extracted with ethyl acetate, the organic layer dried over Na2SO4, and evaporated to yield the title compound as a white foam.
100 mg of the title compound from Step 6 ((S)-CBA-Val-Pec-OH) was dissolved in 3 mL of dichloromethane, and cooled to 0xc2x0 C. via ice water bath. The clear, colorless reaction solution was then treated with 36 mg of 1-hydroxybenzotriazole, 94 mL of diisopropylethylamine, and 170 mg of Asp(OBn)-Pro-Thr(OBn)-propylamide hydrochloride, while stirring. After all solids had dissolved, the reaction solution was treated with 51 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbo-dimide hydrochloride. The solution stirred at 0xc2x0 C. for 1 hour. The reaction flask was then stoppered, sealed with parafilm, and placed in the refrigerator overnight. The dichloromethane solvent was then evaporated and replaced with Ethyl Acetate. The organic solution was washed with water, 1.5M HCl, and 5%Na2CO3, dried over Na2SO4, filtered, and evaporated thoroughly. The entire sample was flash chromatographed eluting with 5% MeOH/CHCl3. The title compound (s-CBA-Val-Pec-Asp(OBz)-Pro-Thr(OBz)-propylamide) was successfully isolated (62 mg) as a white crystalline solid.
(S)-3-phenylbutyric acid was hydrogenated using Rh/C in MeOH at 60 psi and 60xc2x0 C. for 3.2 hours. The reduced product was then converted to the title acid chloride by treating with 1 eq of oxalyl chloride, and a catalytic amount of DMF, in toluene for 90 minutes, followed by evaporation of the volatile materials.
To a room temperature solution of commercially available Boc-Threonine(O-benzyl)-OH (8.00 g, 25.0 mmol) and cesium carbonate (11.32 g, 34.7 mmol) in dry DMF (15.5 mL) under a N2 atmosphere, was added by syringe allyl bromide (3.01 mL, 34.7 mmol). After stirring overnight the mixture was concentrated, diluted with H2O (75 mL) and extracted with ethyl acetate (3xc3x9725 mL). The combined organic layers were washed with saturated NaHCO3 solution (2xc3x9725 mL), H2O (2xc3x9725 mL), and brine (25 mL), then dried with MgSO4. The filtrate was concentrated to a clear, colorless oil (8.76 g). 1H NMR (400 MHz) data was consistent for the Boc compound. The oil was dissolved into 4 M HCl in 1,4-dioxane (100 mL) and stirred at room temperature for 30 minutes. The reaction was concentrated to a clear, yellow oil, triturated several times with Et2O, and concentrated to a oil (7.40 g, 109% crude yield). 1H NMR (400 MHz): consistent with proposed structure.
To a 0xc2x0 C. solution of the title compound from Step 9 (6.80 g, 23.8 mmol) and commercially available Boc-proline-OSu (6.81 g, 21.8 mmol) in CH2Cl2 (70 mL) under a N2 atmosphere, was added diisopropylethylamine (4.32 mL, 24.8 mmol). After 2h at 0xc2x0 C. the solution was washed with water (20 mL), 1.0 N KHSO4 solution (20 mL), saturated NaHCO3 (2xc3x9720 mL), H2O (2xc3x9720 mL) and brine (20 mL). Dried over MgSO4. The filtrate was concentrated and purified by medium pressure chromatography (silica gel, 30% ethyl acetate in hexane) to give a white solid (5.66 g). The solid was dissolved into 4 N HCl in 1,4-dioxane (100 mL) at room temperature. After 1 hour the solution was concentrated, triturated with Et2O several times, and concentrated to give the title compound as a sticky, white solid (4.61 g, 55% yield). 1H NMR (400 MHz): consistent with proposed structure.
To a 0xc2x0 C. solution of the tile compound from Step 10 (4.61 g, 12.0 mmol), Boc-Asp(O-benzyl)-OH (3.89 g, 12.0 mmol), 1-hydroxybenzotriazole (4.41 g, 32.6 mmol) and 4-methyl morpholine (1.39 g, 13.8 mmol) in CH2Cl2 (65 mL) under a N2 atmosphere, was added 1-(3-dimethylaminopropyl)-3-ethyl-carbodiimide hydrochloride (3.28 g, 17.1 mmol). The solution was stirred at 0xc2x0 C. for 1 hour and then room temperature overnight. It was then concentrated and redissolved into CH2Cl2 (100 mL), and washed with 1.0 N KHSO4 solv (2xc3x9725 mL), saturated NaHCO3 solv (2xc3x9725 mL), H2O (2xc3x9725 mL), and brine (25 mL). Dried over MgSO4. The filtrate was concentrated and purified by medium pressure chromatography (silica gel, 30% ethyl acetate in hexane) to give a clear, colorless oil (6.88 g). The oil was dissolved into 4N HCl in dioxane at rt. After 30 minutes the solution was concentrated to a white solid, triturated with Et2O several times, and dried in a 50xc2x0 C. vacuum oven to give the title compound as a white solid (4.65 g, 66% yield). 1H NMR (400 MHz): consistent with proposed structure. Anal. Calcd for C30H38N3O7Cl1: C, 61.27; H, 6.51; N, 7.15. Found: C, 61.05; H, 6.71; N, 7.14.
Commercially available Boc-Lys(Z)-OH was dissolved in dimethylformamide (DMF), and cooled to 0xc2x0 C. via ice water bath. The solution was treated with 1.2 equivalents of 1-hydroxybenzotriazole, 2 equivalents of diisopropylethylamine, and 1.5 equivalents of commercially available pipecolic methyl ester hydrochloride (Pec-OMe.HCl). After all solids were dissolved, the reaction was treated with 1.2 equivalents of 1-(3-dimethyl-aminopropyl)-3-ethylcarbodiimide hydrochloride. Reaction progress was monitored by TLC. After 5 hours, the DMF was thoroughly evaporated. The oily residue was dissolved in water/ethyl acetate. The organic portion was washed with 1.5 M HCl and 5% NaHCO3, dried over MgSO4, and filtered. The solution was evaporated to a clear, colorless oil. The entire sample was then dissolved in 4.0 M HCl in dioxane, stirred for one hour at rt, and thoroughly evaporated to yield the title compound, Lys(Z)-Pec-OMe.HCl, as a white foam.
1.0 g of the title compound from Step 8 (S)-3-cyclohexylbutyroyl chloride was dissolved in 15 mL ethyl acetate, and treated with 1.5 equivalents (eq) of the title compound from Step 12, Lys(Z)-Pec-OMe.HCl, followed by 3 eq of NaHCO3 dissolved in 5 mL of water. After 30 minutes, the organic portion was washed with water, 1.5 M HCl and 5% NaHCO3. The solution was dried over Na2SO4 and evaporated to a clear, colorless oil (89.4%). The entire sample was chromatographed by flash column eluting with 5% MeOH/CHCl3 to remove a baseline impurity. The entire sample was then hydrolyzed using 3 eq of KOH dissolved in water and tetrahydrofuran (THF) as a cosolvent. The reaction stirred vigorously for 5 hours. The THF was evaporated and the aqueous portion was acidified with concentrated HCl. The mixture was extracted with ethyl acetate, dried over Na2SO4, and the solution evaporated to a clear, colorless oil (72%), which was lyophilized from water to give the title compound as a white solid.
The title compound from Step 13 (300 mg), (S)-CBA-Lys(Z)-Pec-OH, was dissolved in 3 mL of CH2Cl2, and cooled to 0xc2x0 C. via ice bath. The clear, colorless reaction solution was then treated with 76 mg 1-hydroxybenzotriazole, 145 xcexcL diisopropylethylamine, and 365 mg of the title compound from Step 11, Asp(OBz)-Pro-Thr(OBz)-allyl ester.HCl, while stirring. After all solids had dissolved, the reaction solution was treated with 108 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide.HCl. After 4.5 hours, the CH2Cl2 was evaporated and replaced with ethyl acetate. The organic solution was washed with water, 1.5 M HCl and 5% NaHCO3, dried over Na2SO4, and evaporated to a clear, colorless oil. The entire sample was chromatographed by flash column eluting with 5% MeOH/CHCl3. The title compound (407 mg), (S)-CBA-Lys(Z)-Pec-Asp(OBn)-Pro-Thr(OBn)-allyl ester, was isolated as a clear, colorless oil (67.5% yield).
2.2 mg of palladium acetate were dissolved in 2 mL dry THF. This clear, light orange solution was then treated with 5 mg of triphenylphosphine. In a separate vial, 138 xcexcL of triethylamine was diluted with 1 mL dry THF and treated with 29 xcexcL of formic acid. This complex was then added to the reaction solution, which promptly turned a dark green color. Finally, the title compound from Step 14 (407 mg), (S)-CBA-Lys(Z)-Pec-Asp(OBn)-Pro-Thr(OBn)-allyl ester was dissolved in 10 mL dry THF and added to the reaction solution. The atmosphere within the reaction vessel was evacuated and replaced with a steady stream of nitrogen. Reaction progress was monitored by TLC. After 86 hours, the solution was filtered through celite, and the clear, pale yellow filtrate was thoroughly evaporated to a sticky foam. The entire sample was chromatographed on silica gel, eluting with 10% MeOH/CHCl3 plus 0.5% HOAc. After purification and lyophilization, 208 mg of the title compound, (S)-CBA-Lys(Z)-Pec-Asp(OBn)-Pro-Thr(OBn)-OH was isolated as a white solid (52.8% yield).
60 mg of the title compound from Step 15, (S)-CBA-Lys(Z)-Pec-Asp(OBz)-Pro-Thr(OBz)-OH were dissolved in 2 mL of CH2Cl2, and cooled to 0xc2x0 C. via ice water bath. The clear, colorless reaction solution was treated with 9 mg of 1-hydroxybenzotriazole, 22 xcexcL of diisopropylethylamine, and 11 xcexcL of commercially available leucinol, while stirring. After all solids had dissolved, the reaction was treated with 13 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide. The reaction flask was then stoppered, sealed with parafilm, and placed in the refrigerator overnight. The CH2Cl2 was thoroughly evaporated and replaced with ethyl acetate. The organic portion was washed with water, 1.5 M HCl, and 5% Na2CO3, dried over Na2SO4, and evaporated to yield 48 mg of the title compound, (S)-CBA-Lys(Z)-Pec-Asp(OBz)-Pro-Thr(OBz)-leucinol as a white solid (73% yield).